JP2015086214A - Prophylactic agents for immune diseases - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a prophylactic agent for immune disease which is ingestible routinely, excellent in safety, and of which ingestion inhibits proliferation of immune cells and elicits high IL-10/IL-12 ratio, to provide food and drinks or nutritional compositions or pharmaceuticals, and to provide a method for screening lactic acid bacteria that inhibit proliferation of immune cells and elicit high IL-10/IL-12 ratio.SOLUTION: As a result of intensive study to find a factor in food materials that shows no cytotoxicity, inhibits proliferation of immune cells and elicits high IL-10/IL-12 production ratio, the inventors have found Lactobacillus helveticus, a lactic acid bacterium, to complete the invention.

Description

The present invention is a preventive agent for immune diseases that can be ingested daily, is excellent in safety, suppresses the proliferation of immune cells by ingestion, and induces a high IL-10 / IL-12 ratio, and its The present invention relates to a screening method. Furthermore, this invention relates to the food-drinks, nutritional composition, and pharmaceutical which mix | blend this immune disease preventive agent.

  When the immune response is appropriately controlled, it works beneficially to the host as a defense function against external enemies, but when the response is excessive, it causes various immune diseases such as enteritis, rheumatism, and allergies. Overproduction of inflammatory cytokines has been pointed out as a cause of such immune diseases.

Therefore, appropriately controlling the production of inflammatory cytokines is effective for the prevention and treatment of immune diseases caused by inflammatory cytokines. In addition, since cytokine production is usually caused by immune cell proliferation, suppression of immune cell proliferation is also considered effective for prevention and treatment of immune diseases caused by inflammatory cytokines. .

Among the indicators of immune status, there is an IL-10 / IL-12 ratio which is a production ratio of IL-10, which is an anti-inflammatory cytokine, and IL-12, which is an inflammatory cytokine. Increasing this IL-10 / IL-12 ratio is believed to be effective against inflammatory diseases.

  As an inhibitor of immune cell proliferation, for example, tacrolimus (Patent Document 1), which is used as an inhibitor of rejection in organ transplantation and as a therapeutic agent for rheumatoid arthritis, and as an enhancer of IL-10 production, cirrhosis and stomatitis Uchinchin-to (Non-patent Document 1), which is a therapeutic agent, is disclosed. However, while drug therapy can be expected to be effective, side effects need to be taken into account.

In recent years, lactic acid bacteria have been used for prevention and treatment of immune diseases. For example, a plurality of lactic acid bacteria to produce increased production of IL-10 / IL-12 ratio in the immune cells in vitro that, be alleviated inflammatory conditions of the animals were developed enterocolitis are reported in in vivo (Non-patent Document 2). In human studies, patients with irritable bowel syndrome have a low IL-10 / IL-12 ratio produced by lymphocytes in the blood, but by taking Bifidobacterium ingants 35624, IL-10 / IL It has been reported that the -12 ratio was increased and symptoms improved. (Non-patent document 3).

Non-Patent Document 4 describes that Lactobacillus acidophilus L-92 is forcibly orally administered to mice to induce apoptosis and suppress the proliferation of T cells, which are a kind of immune cells.

Patent 3-38276

Yamashiki, M. et al., Clinical science, 99 (5): 421-431, 2000 Foligne, B. et al., World J Gastroenterol., 13 (2): 236-243, 2007 O’Mahony, L. et al., Gastroenterology, 128: 541-551, 2005 Kanzato, H. et al., Immunobiology, 213 (5): 399-408, 2008

From the above, it can be seen that increasing the IL-10 / IL-12 ratio and suppressing the proliferation of immune cells are effective for the prevention and treatment of immune diseases. However, these two actions are considered to be opposite to each other. Usually, when the production of cytokines such as IL-10 and IL-12 increases, immune cell activation is accompanied by proliferation of immune cells. Therefore, it is considered that cytokine production is suppressed when the proliferation of immune cells is suppressed. In existing reports, only one of cytokine production promotion effects such as suppression of proliferation of immune cells or increase of IL-10 / IL-12 production ratio has been reported, and these two actions are expressed simultaneously. There is no known substance that can be used, especially lactic acid bacteria.

The present invention is a preventive agent for immune diseases that can be ingested daily, is excellent in safety, and suppresses the proliferation of immune cells and induces a high IL-10 / IL-12 ratio by ingestion, and It is an object to provide a food or drink, a nutritional composition, or a pharmaceutical product that has been given this function.

It is another object of the present invention to provide a screening method for lactic acid bacteria that, when ingested, suppresses the proliferation of immune cells and induces a high IL-10 / IL-12 ratio.

The present invention is an invention having any of the following configurations.
(1)
An immune disease preventive agent comprising, as an active ingredient, a lactic acid bacterium belonging to Lactobacillus helveticus .
(2)
The lactic acid bacterium is Lactobacillus helveticus SBT2171 strain, the immune disease preventive agent according to (1),
(3)
The lactic acid bacterium is Lactobacillus helveticus SBT2161 strain, the immune disease preventive agent according to (1).
(4)
(1) thru | or the immune disease preventive agent in any one of (3), The food-drinks for immune disease preventive agents characterized by the above-mentioned, the nutrition composition for immune disease preventive agents, or the pharmaceutical for immune disease preventive agents.
(5)
An agent for suppressing immune cell proliferation and increasing the IL-10 / IL-12 production ratio, comprising lactic acid bacteria belonging to Lactobacillus helveticus as an active ingredient.
(6)
The said lactic acid bacterium is Lactobacillus helveticus ( Lactobacillus helveticus ) SBT2171 strain | stump | stock, The immune cell proliferation suppression and IL-10 / IL-12 production ratio increase agent of Claim 5 characterized by the above-mentioned.
(7)
The said lactic acid bacterium is Lactobacillus helveticus ( Lactobacillus helveticus ) SBT2161 strain | stump | stock, The immune cell proliferation suppression and IL-10 / IL-12 production ratio increase agent of Claim 5 characterized by the above-mentioned.
(8)
(5) to (7) an immune disease growth-preventing agent and an IL10 / IL12 production ratio increasing food or drink, an immune cell growth inhibition and IL10 / IL12 production ratio, characterized by comprising the immune disease prevention agent according to any one of A nutritional composition for an increasing agent or a pharmaceutical agent for suppressing immune cell proliferation and increasing an IL-10 / IL-12 production ratio.
(9)
The screening method of lactic acid bacteria characterized by including the following step at least.
(A) screening for lactic acid bacteria that increase the ratio of IL-10 production / IL-12 production produced from immune cells.
(B) A step of screening for lactic acid bacteria that suppress cell proliferation of immune cells.

Lactobacillus helveticus Lactobacillus helveticus is an immune disease preventive agent that can suppress the proliferation of immune cells and induce a high IL-10 / IL-12 ratio. It is effective in preventing or treating immune diseases such as rheumatism and hepatitis.

It is the figure which showed the proliferation activity of the immune cell of the mouse | mouth which added lactic acid bacteria. It is the figure which showed the IL-10 production amount of the immune cell of a mouse | mouth which added lactic acid bacteria. It is the figure which showed the IL-12 production amount of the immune cell of a mouse | mouth which added lactic acid bacteria. It is the figure which showed IL-10 / IL-12 production ratio computed from the IL-10 production amount of a mouse | mouth immune cell which added lactic acid bacteria, and IL-12 production amount. It is the figure which showed IL-10 / IL-12 production ratio computed from the IL-10 production amount of a mouse | mouth immune cell which added lactic acid bacteria, and IL-12 production amount. It is the figure which showed the number of living cells of the mouse | mouth immune cell which added lactic acid bacteria.

In the present invention, a strain of Lactobacillus helveticus having an immune disease preventing action that suppresses the proliferation of immune cells and induces a high IL-10 / IL-12 ratio is used. The type of Lactobacillus helveticus is not limited, but examples include Lactobacillus helveticus SBT2171 strain (FERM BP-5445), Lactobacillus helveticus SBT2161 strain (NITE ABP-01707), Lactobacillus helveticus SBT0621 strain (FERMP-21883) and Lactobacillus helveticus SBT 0064 strain (FERMP-21079). In particular, Lactobacillus helveticus SBT 2 1 71 1 strain has been reported as a strain of the genus Lactobacillus having high proteolytic activity without causing bitterness, odor and the like (Japanese Patent Laid-Open No. 7-2 7 4 9 4 9).

  Bacterial cells of Lactobacillus helveticus can be used as an active ingredient of the present invention as they are after culturing according to a conventional method of lactic acid bacteria culture and separated from the resulting culture by means of collecting bacteria such as centrifugation. Not only cells that have been purely isolated as cells, but also cultures, suspensions, other cell-containing materials, and cytoplasm and cell wall fractions obtained by treating cells with enzymes or physical means Can do. Furthermore, in formulation, excipients, stabilizers, flavoring agents, and the like that are permitted in the formulation are mixed as appropriate and concentrated, freeze-dried, or dried by heating to form dead cells. These dried products, concentrates, and pastes are also included. In addition, as long as it does not interfere with the immune disease preventive action of Lactobacillus helveticus cells, an excipient, binder, disintegrant, lubricant, flavoring agent, suspending agent, coating agent, and other optional agents can be added. It can also be formulated by mixing. The dosage form can be tablets, capsules, granules, powders, powders, syrups, etc., and these are preferably administered orally.

Since the active ingredient of the present invention can achieve an immune disease preventive action by oral administration, it is used as a food and drink for immune disease prevention, a nutritional composition for preventing immune disease, and a pharmaceutical for preventing immune disease. Can do. The form of the food and drink for immune disease prevention may be any form as long as it does not interfere with the immune disease preventive action, and the lactic acid bacteria itself, fermented milk obtained by culturing the lactic acid bacteria and the cheese itself Furthermore, these cells, fermented milk, cheese, etc. may be used as materials, and may be made into bread, snacks, cakes, puddings, beverages, fermented milk, noodles, sausages and other foods and drinks, and various powdered milk In addition, it is also possible to mix | blend with infant food, a nutritional composition, etc.

In addition, the immune disease preventive agent of the present invention is blended, and the immune disease preventive agent, the immune disease preventive food and drink, the immune disease preventive nutrition composition, the immune disease preventive drug, and the like, or the processed product of these materials In the case where it is used, the content ratio of the lactic acid bacteria is not particularly limited, and may be appropriately adjusted according to the ease of production, the preferable daily dose, and the like. For example, when the dosage form is liquid, it is preferably 1 × 10 ⁵ cells / ml to 1 × 10 ¹⁰ cells / ml, and when it is solid, 1 × 10 ⁵ cells / g to 1 × 10 0 ¹⁰ cells / g is preferable.

Since these immune disease preventive agents, immune disease preventive foods and drinks, immune disease preventive nutritional compositions, and immune disease preventive pharmaceuticals have the ability to prevent immune diseases, they have various pathological conditions caused by the above-mentioned immune disorders. It can be very useful for prevention, treatment, improvement and prevention of recurrence.
In order to exert the ability to prevent immune diseases of the present invention, in the case of an adult, it is desirable to ingest 1-1,000 mg by weight of lactic acid bacteria. Lactic acid bacteria have been used in the manufacture of fermented milk and cheese since ancient times, and the immune disease preventive agent, immune disease preventive food and drink, immune disease preventive nutrition composition, and immune disease preventive drug of the present invention are safe. There is a feature that there is no problem.

The screening method of the present invention is a method for screening lactic acid bacteria that suppress the proliferation of immune cells from various lactic acid bacteria, and a method for screening lactic acid bacteria that induce a high IL-10 / IL-12 ratio among various lactic acid bacteria. Is a combination.

Hereinafter, the present invention will be described in more detail with reference to examples. However, these are merely illustrative, and the present invention is not limited thereto.

(Measurement of immune cell proliferation activity)
As described below, in vitro , 43 types of lactic acid bacteria shown in Table 1 were added to mouse immune cells and cultured, and their effects on the proliferation activity of immune cells were evaluated and compared with controls (without addition of lactic acid bacteria) Thus, lactic acid bacteria that reduce the growth activity were screened.

(Preparation of lactic acid bacteria)
The lactic acid strains used in the test are shown in Table 1. Lactobacillus helveticus ( Lactobacillus helveticus ) strain SBT2171, Streptococcus thermophilus and Lactobacillus (excluding Lactobacillus kefiranofaciens and Lactobacillus plantarum ) were cultured in MRS medium (Difco Laboratories, Detroit, Mich.) For 16 hours. Among the Lactobacillus Lactobacillus kefiranofaciens, Lactobacillus plantarum, and Lactococcus lactis were cultured 30 ° C. 16 hours in MRS medium. Bifidobacterium was cultured at 37 ° C. for 16 hours in a GAM medium (Nissui Seiyaku Co., Tokyo, Japan) supplemented with 1% glucose. Leuconostoc was cultured at 30 ° C. for 16 hours in a GAM medium supplemented with 1% glucose.
The preparation of the bacterial cell sample from the culture solution was performed according to a conventional method. Specifically, after culturing each lactic acid strain in Table 1 according to the above procedure, the cells were collected from each lactic acid bacteria culture solution by centrifugation, washed with sterile ultrapure water, and then lyophilized. The lyophilized powder was suspended in sterile ultrapure water at a concentration of 10 mg / ml. The bacterial suspension was heated at 80 ° C. for 30 minutes to obtain a test sample. Cryopreserved until used for testing.

ssp.; subspecies
Lb.: Lactobacillus, Lc.: Lactococcus, B.: Bifidobacterium, Leu.: Leuconostoc
T: Type strain（基準株）
JCM: JapanCollection of Microorganisms, Saitama, Japan.
ATCC: American type culture collection, Manassas, USA.
NCFB: National Collection for Food Bacteria, Reading, UK.

ssp .; subspecies
Lb .: Lactobacillus, Lc .: Lactococcus, B .: Bifidobacterium, Leu .: Leuconostoc
T: Type strain
JCM: Japan Collection of Microorganisms, Saitama, Japan.
ATCC: American type culture collection, Manassas, USA.
NCFB: National Collection for Food Bacteria, Reading, UK.

Preparation of mouse immune cells
RPMI 1640 medium (Life Technologies, Gaithersburg, MD) with 10% inactivated fetal bovine serum (guaranteed endotoxin) (Life Technologies), 10 mM HEPES (Life Technologies), 2 mM L-glutamine (Life Technologies), 100 U RPMI medium was prepared by adding / mL penicillin (Life Technologies), 100 μg / mL streptomycin (Life Technologies) and 0.05 mM 2-mercaptoethanol. Mesenteric lymph nodes were removed from C57BL / 6J mice (7-10 weeks old, male; Charles River, Japan), added to RPMI medium and ground, and 70 μm-cell strainer (BD Biosciences, San Jose, Filtered with CA) to obtain a cell dispersion. The cells in the dispersion were washed twice with RPMI medium and then redispersed in RPMI medium.

Cell proliferation activity measurement The prepared cells were seeded at a density of 5.0 × 10 ⁴ cells / well in a 96-well round-bottomed plate (iwaki, Japan), and the same number of anti-mouse CD3 / CD28 antibody beads (Life Technologies) as the number of cells, Each lactic acid bacteria sample (final concentration 10 μg / ml) was added to 200 μl, and the cells were cultured for 3 days at 37 ° C. and 5% CO ₂ . PrestoBlue (R) Cell Viability Reagent (Life Technologies), a reagent that emits intense fluorescence, was added to each well according to the protocol of the product as the proliferation activity of the cells increased after 3 days of culture. After further culturing for 7 hours, the fluorescence intensity of the culture supernatant was measured. The fluorescence intensity to which each lactic acid strain was added was shown in FIG. 1 as a relative value with the fluorescence intensity of the control group cultured by adding only anti-mouse CD3 / CD28 antibody beads to the cells as 100.

As shown in FIG. 1, all five strains of Lactobacillus helveticus (SBT2161, SBT0064, SBT0621, SBT2171, and JCM1120 ^T ) had lower fluorescence intensity than the control. That is, when Lactobacillus helveticus ( Lactobacillus helveticus ) was added to a culture system, it turned out that the proliferation activity of an immune cell reduces.

(Measurement of IL-10 / IL-12 production ratio (1))
Among the lactic acid bacteria in Table 1, Lactobacillus gasseri JCM1131 ^T strain whose cell proliferation activity was increased compared to the control, Lactobacillus salivarius ssp. Salivarius SBT11459, Lactobacillus plantarum SBT11457, and cell proliferation whose cell proliferation activity was almost the same as the control A total of 4 strains of Lactobacillus helveticus SBT2171 with reduced activity were selected and subjected to measurement of IL-10 / IL-12 production ratio. As described below, in vitro , the above four strains were added to mouse immune cells, cultured, and screened for lactic acid bacteria that increased the IL-10 / IL-12 production ratio compared to controls (without addition of lactic acid bacteria). .

Preparation of mouse immune cells Preparation was carried out in the same manner as in the above-mentioned measurement of cell proliferation activity.

Table 2 shows the lactic acid bacteria used in the preparation test of lactic acid bacteria. Preparation of the cell sample from the culture broth was based on the method of Foligne et al. (Non-patent Document 2). Specifically, each lactic acid strain was cultured in MRS medium (Difco Laboratories) at 37 ° C. for 16 hours, and the cells were collected from each lactic acid bacteria culture solution by centrifugation. After washing with PBS (−), it was suspended in PBS (−) containing 20% glycerol at a concentration of 1 × 10 ⁹ cfu / ml. Cryopreserved at −80 ° C. until used for testing.

Measurement of IL-10 / IL-12 production ratio
Cells prepared at a concentration of 2.0 × 10 ⁶ cells / ml are seeded in a 24-well plate (iwaki) 1 ml per well (2.0 × 10 ⁶ cells / well) and adjusted to a concentration of 1 × 10 ⁹ cfu / ml 20 μl of each lactic acid bacteria sample was added (2.0 × 10 ⁷ cfu / well). In the control group, 20 μl of PBS (−) containing 20% glycerol was added instead of the lactic acid bacteria sample. The cells were cultured for 24 hours under conditions of 37 ° C. and 5% CO ₂ , and after culturing, the whole culture solution containing the cells was centrifuged to obtain a culture supernatant. IL-10 production in the supernatant was measured using Mouse IL-10 Ready-SET-Go! (EBioscience, San Diego, Calif.) Of an IL-10 measurement kit, and the numerical values are shown in FIG. . Similarly, IL-12 production in the supernatant was measured using mouse IL-12 p70 (R & D Systems, Minneapolis, Minn.) Of IL-12 measurement kit, and the numerical values are shown in FIG.

From FIG. 2, Lactobacillus helveticus SBT2171 strain strongly induced IL-10 production from immune cells. The effect was similar to that of Lb. salivarius ssp. Salivarius, which is reported to have a large production amount in Non-Patent Document 2, and was stronger than other lactic acid bacteria. As shown in FIG. 3, with regard to IL-12, Lactobacillus helveticus SBT2171 strain has a weaker production promoting effect than other lactic acid bacteria, and its production amount is similar to that of the control group to which no cells were added. It was. The IL-10 / IL-12 ratio calculated from these IL-10 production and IL-12 production is shown in FIG. FIG. 4 showed that the Lactobacillus helveticus SBT2171 strain was the highest among all groups including the control group. That is, it was found that Lactobacillus helveticus induces a high IL-10 / IL-12 ratio.

As a result of the above-described measurement of immune cell proliferation activity and measurement of IL-10 / IL-12 production ratio, Lactobacillus as a lactic acid bacterium that suppresses the proliferation of immune cells and induces a high IL-10 / IL-12 ratio. Helveticas ( Lactobacillus helveticus ), especially Lactobacillus helveticus SBT2171 strain could be screened.

(Measurement of IL-10 / IL-12 production ratio (2))
Among the lactic acid bacteria in Table 1, Lactobacillus helveticus SBT2161 strain (NITE ABP-01707), Lactobacillus helveticus SBT0621 strain (FERMP-21883), Lactobacillus helveticus SBT 0064 strain (FERMP-21079) A total of 3 strains were selected and subjected to measurement of IL-10 / IL-12 production ratio. As described below, in vitro , the above four strains were added to mouse immune cells, cultured, and screened for lactic acid bacteria that increased the IL-10 / IL-12 production ratio compared to controls (without addition of lactic acid bacteria). .

Preparation of mouse immune cells Adjustment was made in the same manner as in the measurement of the IL-10 / IL-12 production ratio (1) described above.

Preparation of lactic acid bacteria Lactic acid strains used in the test are shown in Table 3. The bacterial cell sample from the culture solution was prepared by the same method as the above-described measurement of IL-10 / IL-12 production ratio (1).

Measurement of IL-10 / IL-12 production ratio The measurement was performed in the same manner as in the measurement of IL-10 / IL-12 production ratio (1) described above. FIG. 5 shows the IL-10 / IL-12 ratio calculated from the IL-10 production amount and the IL-12 production amount.

From FIG. 5, the Lactobacillus helveticus SBT2161 strain (NITE ABP-01707), the Lactobacillus helveticus SBT0621 strain (FERMP-21883), and the Lactobacillus helveticus SBT 0064 strain (FERMP-21079) have no added cells. The IL-10 / IL-12 ratio was induced significantly higher than that of the group. That is, it was found that Lactobacillus helveticus induces a high IL-10 / IL-12 ratio.

As a result of the above-described measurement of immune cell proliferation activity and measurement of IL-10 / IL-12 production ratio, Lactobacillus as a lactic acid bacterium that suppresses the proliferation of immune cells and induces a high IL-10 / IL-12 ratio. Helveticas ( Lactobacillus helveticus ), especially Lactobacillus helveticus strain SBT2161 (NITE ABP-01707) could be screened.

(Cytotoxicity measurement)
Usually, when the production of cytokines such as IL-10 and IL-12 increases, immune cell activation is accompanied by proliferation of immune cells. However, the above results indicate that Lactobacillus helveticus induces a high IL-10 / IL-12 ratio and suppresses the proliferation of immune cells. In order to investigate the cause, Lactobacillus gasseri strain JCM1131 and Lactobacillus helveticus were selected, and cytotoxicity was evaluated in vitro .

Preparation of mouse immune cells Preparation was carried out in the same manner as in the above-mentioned measurement of cell proliferation activity.

Table 3 shows the lactic acid strains used in the lactic acid bacteria preparation test. The culture conditions for each lactic acid bacterium and the method for preparing the test sample are the same as in the above-described cell proliferation activity measurement.

Cytotoxicity measurement Prepared cells are seeded at a density of 5.0 × 10 ⁴ cells / well in a 96-well round-bottomed plate (iwaki), and each lactic acid bacteria sample (final concentration 10 μg / ml) is added to 200 μl to 37 ° C and 5% CO ₂ were cultured for 3 days. After culturing for 3 days, CyQUANT® Direct Cell Proliferation Assay kit (Life Technologies) was added according to the protocol of the product, further cultured for 1 hour, and fluorescence intensity was measured. The number of living cells calculated from the fluorescence intensity is shown in FIG.

From FIG. 6, the number of living cells when Lactobacillus helveticus ( Lactobacillus helveticus ) SBT2171 was added was not significantly different from the case where lactic acid bacteria were not added (control group). That is, it was shown that this strain does not affect cell survival when there is no growth stimulation such as antibody beads, that is, this strain has no obvious cytotoxicity. Therefore, Lactobacillus helveticus induces a high IL-10 / IL-12 ratio and can suppress the proliferation of immune cells.

(Manufacturing fermented milk)
Lactobacillus helveticus ( Lactobacillus helveticus ) SBT2171 strain cultured in 100 g of skim milk at 37 ° C. for 12 hours was inoculated into 3 kg of the same medium and cultured at 37 ° C. for 12 hours. The whole amount of fermented milk after the culture was used as a starter, and 100 kg of skim milk was fermented at 32 ° C. for 20 hours to obtain fermented milk of Lactobacillus helveticus SBT2171 strain having an immune disease preventing action. The viable count of Lactobacillus helveticus SBT2171 was 8.2 × 10 ⁸ cells / g.

(Manufacture of powdered tablets of SBT2171 strain)
1 part of the cell powder obtained in (0024) of Example 1 was mixed with 4 parts of skim milk powder, and this mixed powder was tableted 1 g by a conventional method with a tableting machine, and Lactobacillus helveticus ( Lactobacillus helveticus). ) A cell powder tablet having an immune disease-preventing action, containing 200 mg of SBT2171 strain cells was obtained.

(Manufacture of capsules)
The raw materials were mixed according to the formulation shown in Table 5, granulated, and then filled into capsules to produce capsules that were given immune disease prevention production.

(Cheese production)
Several kinds of low-fat hard natural cheeses were produced using raw milk adjusted so that the fat percentage in cheese was 12 to 30%. That is, the raw milk with adjusted fat percentage was sterilized by heating at 75 ° C. for 15 seconds, then cooled to 30 ° C., and 0.01% calcium chloride was added. Next, 0.7% of commercially available lactic acid bacteria starter (manufactured by Christian Hansen) and 1% of Lactobacillus helveticus JCM1120 ^T strain (reference strain) are added to these raw milks, and 0.003% of rennet is added. After coagulating the milk, it is cut and the pH is 6.2.
The mixture was stirred until ˜6.1, and whey was discharged to obtain curd grains. Furthermore, this curd grain was mold-packed and squeezed and further salted to produce gouda cheese type low-fat hard natural cheese.

Conventionally, in order to suppress the proliferation of immune cells and increase the IL-10 / IL-12 ratio, two or more types of lactic acid bacteria were required. However, one type of Lactobacillus helveticus of the present invention was used. The above-mentioned effects can be obtained with lactic acid bacteria. By suppressing the proliferation of immune cells, the rejection of organ transplants and the therapeutic effect of rheumatoid arthritis, as well as the treatment of cirrhosis and stomatitis by increasing the IL-10 / IL-12 ratio The effects can be expected together, and further, the possibility of being effective for new immune diseases can be expected by having these effects together.

Claims (9)

An immune disease preventive agent comprising, as an active ingredient, a lactic acid bacterium belonging to Lactobacillus helveticus . 2. The preventive agent for immune diseases according to claim 1, wherein the lactic acid bacterium is Lactobacillus helveticus SBT2171 strain. The lactic acid bacterium is Lactobacillus helveticus SBT2161 strain, the immune disease preventive agent according to claim 1, wherein the lactic acid bacterium is Lactobacillus helveticus strain SBT2161. A food or drink for an immune disease preventive, a nutritional composition for an immune disease preventive, or a pharmaceutical for an immune disease preventive comprising the immune disease preventive agent according to any one of claims 1 to 3. An agent for suppressing immune cell proliferation and increasing the IL-10 / IL-12 production ratio, comprising lactic acid bacteria belonging to Lactobacillus helveticus as an active ingredient. The said lactic acid bacterium is Lactobacillus helveticus ( Lactobacillus helveticus ) SBT2171 strain | stump | stock, The immune cell proliferation suppression and IL-10 / IL-12 production ratio increase agent of Claim 5 characterized by the above-mentioned. The said lactic acid bacterium is Lactobacillus helveticus ( Lactobacillus helveticus ) SBT2161 strain | stump | stock, The immune cell proliferation suppression and IL-10 / IL-12 production ratio increase agent of Claim 5 characterized by the above-mentioned. A food / drink for immune cell growth suppression and IL10 / IL12 production ratio increasing agent, comprising an immune disease prevention agent according to any one of claims 5 to 7, an immune cell proliferation suppression and IL10 / IL12 production ratio increase agent Nutritional composition, or pharmaceutical agent for suppressing immune cell proliferation and increasing IL-10 / IL-12 production ratio. A screening method for lactic acid bacteria, comprising at least the following steps: (A) a step of screening for lactic acid bacteria that suppress cell growth of immune cells.
(B) screening for lactic acid bacteria that increase the ratio of IL-10 production / IL-12 production produced from immune cells.

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